The focus of this proposal is the muscle nicotinic acetylcholine receptor (AChR), the neurotransmitter-gated ion channel that mediates fast synaptic transmission at the vertebrate neuromuscular junction (NMJ). The broad objective of this proposal is to understand how the structure of this receptor-channel gives rise to its function, and how this function is exquisitely tailored to the physiological needs of the vertebrate NMJ. Throughout the years, research on the AChR has provided answers to fundamental questions on the physics, chemistry, and biology of ion channels that cannot be addressed with (probably) any other channel. Our main tools are single- channel and macroscopic electrophysiology (using both in-vitro and in-situ approaches), protein engineering, and quantitative thinking. Our four Specific Aims are: I) To determine the location and physical nature of the AChR's gate(s), and its rearrangement upon opening and shutting of the channel, II) To understand the relationship between structure and function in the AChR's selectivity filter, and the coupling between ion- occupancy and gating/desensitization, III) To estimate the value of the wild-type AChR's unliganded-gating equilibrium constant, and IV) To define the contribution of desensitization to the synaptic response of the AChR. It is worth noting that, in addition to their obvious relevance in the relay of neuron-to-muscle signals, ACh, the AChR, and the NMJ have served as models of neurotransmitters, receptors, and synapses, in general. Hence, the knowledge derived from the proposed experiments is poised to have a broad impact on our basic understanding of postsynaptic receptors and fast synaptic transmission. It should also be emphasized that AChR-mediated neurotransmission supports not only motor but, also, autonomic and cognitive function and, therefore, that it is compromised not only in myasthenias but, also, in cardiovascular diseases and neurodegenerative disorders such as Alzheimer disease. Furthermore, AChRs are also expressed in the airways, keratinocytes, lymphocytes, and endothelial cells where they are mediators of cell differentiation, proliferation, and resistance to apoptosis and, thus, of cancer. Indeed, it is becoming increasingly clear that nicotine addiction (which is caused by the interaction of nicotine with AChRs in behavior- controlling areas of the brain) increases the risk of developing lung and oral cancer owing to the excessive activation of these non-muscle/non-neuronal AChRs by inhaled nicotine and nicotine-derived nitrosamines while smoking.